The long-term objective of my research is to develop and apply combinatorial chemistry for basic research and drug development. Protein kinases represent one of the largest protein superfamilies. These enzymes catalyze the transfer of gamma-phosphate of ATP (or GTP) to the protein alcohol groups (on Ser or Thr) and/or protein phenolic groups (in Tyr). Over 200 different superfamily member had been recognized from mammalian sources alone. Many of the enzymes from this superfamily have been proven to play a crucial role in many cell regulatory processes and disease-states. A technology that enables one to rapidly compare the protein kinase profile of normal and disease-tissue may provide critical information in the understanding of the disease process, and enable one to identify relevant target(s) for the development of drugs against the disease. The main objective of this proposal is to develop such a technology using tumor cell lines as a model system. We hypothesize that by coupling the "one-bead one-compound" combinatorial library method with the differential display concept, we can develop a new method to rapidly identify peptide substrates for novel protein kinase activity in disease-tissue. We further hypothesize that using the identified substrate motif, we may be able to identify the deranged enzymes as well as design specific drugs to inhibit them. This method involves the use of the well-established on-bead functional assay of the "one-bead one-compound" combinatorial library method to identify novel peptide substrates for protein kinases. 3T3 mouse fibroblast and v-src transfected 3T3 mouse fibroblasts will be used as a model system to work out this proposed technology. This technology will then be applied to human B-cell lymphoma/peripheral B-lymphocytes, human breast cancer/surrounding normal breast tissue, and human colon cancer/surrounding normal colonic tissue.